An Engineered Split Intein for Photoactivated Protein Trans-Splicing |
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| Authors: | Stanley Wong Abdullah A. Mosabbir Kevin Truong |
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| Affiliation: | 1. Institute of Biomaterials and Biomedical Engineering, University of Toronto, 164 College Street, Toronto, Ontario, M5S 3G9, Canada.; 2. Edward S. Rogers Sr. Department of Electrical and Computer Engineering, University of Toronto, 10 King’s College Circle, Toronto, Ontario, M5S 3G4, Canada.; Imperial College London, UNITED KINGDOM, |
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| Abstract: | Protein splicing is mediated by inteins that auto-catalytically join two separated protein fragments with a peptide bond. Here we engineered a genetically encoded synthetic photoactivatable intein (named LOVInC), by using the light-sensitive LOV2 domain from Avena sativa as a switch to modulate the splicing activity of the split DnaE intein from Nostoc punctiforme. Periodic blue light illumination of LOVInC induced protein splicing activity in mammalian cells. To demonstrate the broad applicability of LOVInC, synthetic protein systems were engineered for the light-induced reassembly of several target proteins such as fluorescent protein markers, a dominant positive mutant of RhoA, caspase-7, and the genetically encoded Ca2+ indicator GCaMP2. Spatial precision of LOVInC was demonstrated by targeting activity to specific mammalian cells. Thus, LOVInC can serve as a general platform for engineering light-based control for modulating the activity of many different proteins. |
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